A study of African swine fever virus in Regional VI of the Disease Investigation Center of Denpasar Bali in Indonesia.

Background and Aims: African swine fever (ASF) is a highly contagious viral disease that causes major economic losses due to morbidity and fatality rates of up to 100% in wild boar and domestic pigs. The disease emerged in Africa in 1921 and then entered several European countries by 1957. In Indonesia, the first outbreak of ASF in 2019 in North Sumatra killed thousands of pigs and quickly spread to 10 out of 34 pig-producing provinces, including Bali and Eastern Nusa Tenggara. As no commercial ASF vaccine is available, the disease has become endemic and continues killing pigs. This study aimed to investigate the epidemiological and virological studies of ASF virus (ASFV) conducted in 2020 and 2021 by the Disease Investigation Center Regional VI of Denpasar Bali, which covers three provinces in Indonesia, including Bali, Western Nusa Tenggara, and Eastern Nusa Tenggara. Materials and Methods: A total of 5402 blood samples were sent to the laboratory to detect ASFV infection using quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay tests. The virological studies were performed by culturing local ASFV isolates obtained from field cases in primary macrophages and confirmation of viral growth by qPCR. Results: The qPCR results show that only 156/4528 (3.4%) of samples originating from Bali and Eastern Nusa Tenggara were ASFV-positive with cycle threshold value of 18 to 23, while the virus was not detected in Western Nusa Tenggara. Of 874 serum samples tested, 114 (13%) were antibody positive and were all collected from the two ASFV-affected provinces in 2020. A Bali ASFV isolate (BL21) was isolated and characterized molecularly. Conclusion: These findings suggest that during the time of sampling, ASFV was detected only in Bali and East Nusa Tenggara but not in Western Nusa Tenggara. These findings support the symptomology of ASFV reported in the two regions. Moreover, BL21 may be useful for developing subculture-attenuated vaccines using commercial cell lines. However, the current study has some limitations namely the investigation was not performed during the initial outbreak and no pathological examination of internal organs was conducted.

Assessment of Cell Cycle and Induction of Apoptosis by Shiga-like Toxin Produced by Escherichia coli O157:H7 in T47D Breast Cancer Cells Using Flow Cytometry.

BACKGROUND: The low general toxicity against tumors expressing globotriaosylceramide (Gb3) and Shiga-like toxins produced by E. coli have been proposed as an anti-cancer therapy because of their specific target. This study aimed to determine the potency of the local strains of E. coli O157:H7 isolated from humans and cattle as a new breast cancer therapy by analyzing the cell cycle's inhibition and apoptosis induction. MATERIAL AND METHODS: Approximately 10 cultured T47D cells were subjected to Shiga-like toxin produced by four local isolates of E. coli O157:H7, including KL-48 (2) from humans, and SM-25 (1), SM-7 (1), DS-21 (4) from cattle. Using ATCC 43894 as a control, the treatment was observed for 24 h by two replications. In addition, a FITC-Annexin V and PI assay were used to observe apoptosis and necrosis effect, as well as to analyze the cell cycle using propidium iodide (PI) staining. RESULTS: The results showed the toxicity effect of Shiga in the human T47 D cells line. The viability of the cells is subjected to Shiga-like toxins produced by KL-48 (2), SM7 (1), ATCC 43894, SM-25 (1), and DS-21 (4) isolates decreased with 15.20, 16.36, 22.17,  22.64, and 33.86%, in contrary to control of 94.36%. These were supported by the cells entering the late apoptosis of the cell cycle through each isolate with 67.66, 62.60, 63.68, 63.90, and 54.74%, and a control of 0.01%. Also, the necrosis cell for each treatment of 12.73, 19.3, 10.84, 10.53, and 4.86% was higher than the control of 5.51%. These were confirmed by the higher percentage of the cells treated with toxins of KL-48 (2), SM7(1), ATCC 43894, SM-25 (1), and DS-21 (4), which entered G0-G1 of the cell cycle phase with 66.41, 63.37, 61.52, 55.36, and 47.28%, respectively, than control of 40.69%. Additionally, the toxicity effect was supported by an increase in the cells entering the S and the G2-M phase of the cycle for each treatment. CONCLUSION: It is concluded that the Shiga-like toxin produced by E. coli O157:H7 local isolates can be developed as a drug against breast cancer based on its effect to arrest induction of the cell cycle and inducing apoptosis.

Molecular identification of lactic acid bacteria SR6 strain and evaluation of its activity as an anticancer in T47D cell line.

Background and Aim: Breast cancer is the most common type of cancer in women because it attacks the productive age. Preliminary studies showed that lactic acid bacteria (LAB) strain SR6 from the Bali cattle colon has the potential to act as a superior probiotic. It is also assumed that its bacteriocin structure is specific and has a strong relationship with the specificity of the ligand and its biological activity at a receptor. Therefore, this study aims to assess the use of local LAB strains, which produce bacteriocins as anticancer agents, as well as to identify the bacteria as potent producers molecularly. Materials and Methods: The study was initiated by cultivating LAB SR6 strain from stock isolates on De Man, Rogosa, and Sharpe (Oxoid, CM 0369, England) broth media. It was then confirmed molecularly through analysis of the 16S ribosomal ribonucleic acid gene. Subsequently, its anticancer activity was tested by assessing the cytotoxic activity in T47D cell culture using the 3-(4, 5 dimetiltiazol-2-yl)-2.5-diphenyl tetrazolium bromide (Invitrogen M6494, US) method. Results: The results showed that the LAB strain SR6 was identified molecularly as Pediococcus pentosaceus. Furthermore, it had a toxic effect on T47D cells, which was indicated by the number of deaths after treatment with the extracellular protein of the strain, especially at the 50% total cell volume level. Conclusion: Based on the toxic effect of the strain on human T47D cells, the LAB SR6 isolate, which was identified as P. pentosaceus has the potential to be developed as a good anticancer drug against breast cancer. However, there is a need to carry out an integrated study to fully explore the suitability of bacteriocins as in vivo therapeutics against the disease completely.

Evaluation of Community-Based Dog Welfare and Rabies Project in Sanur, a Sub-district of the Indonesian Island Province of Bali.

The Indonesian island province of Bali experienced its first rabies incursion in 2008. Mass vaccination of the dog population has proven effective and rabies cases in dogs and people have decreased, however the virus is still circulating among the dog population. Vaccination coverage must be maintained until rabies elimination. Increasing efficiency and effectiveness of vaccination campaigns is therefore desired. Community engagement leading to preventative health actions by community members can reduce disease incidence and costs of control. Here we evaluate 2 years of a novel community-based dog welfare and rabies control project (Program Dharma) in the Sanur sub-district. The project engaged the services of people living in the project area with an interest or experience in dogs or community health services. These people spoke with owners within their own community about dog welfare and health, monitored owned and unowned dogs and increased owner and carer efforts to access vaccination and further veterinary services. The evaluation focused on a sample of dogs whose owners had been regularly engaged with project. Vaccination coverage was increased and there were no dog or human rabies cases reported in the project area; the percentage of the dogs that had never been vaccinated was reduced by an average 28.3% (baseline unvaccinated 41-49%, post-project unvaccinated 11-19%). The welfare of dogs improved from an average of 20.7% of dogs with visible welfare problems at baseline to 2.7% after project implementation. Roaming dog density observed on street surveys also decreased in all project areas (24-47% reduction dependent on desa). A participatory evaluation event with a sample of Program Dharma community-based agents highlighted several additional successes, including that the community appeared to welcome and value their services and were beginning to support the cost of project activities. Conversely, challenges included identifying dogs in the database during revisits, sustaining the costs of community member time spent working on Program Dharma activities and the costs of veterinary care, whilst avoiding dependency of owners on free veterinary services. The benefits revealed by the evaluation were judged to be sufficient to extend Program Dharma to new areas, whilst evolving activities to resolve challenges.

Dog Ecology and Rabies Knowledge of Owners and Non-Owners in Sanur, A Sub-District of the Indonesian Island Province of Bali.

This study gained an understanding of dog demographics, owner behaviour, and knowledge, attitudes and practices relating to rabies in three villages in Sanur, Bali, providing insights for an intervention to improve responsible dog ownership. A combination of a census of all dogs, street surveys of roaming dogs, and a Knowledge, Attitude and Practices (KAP) survey was used to study both dogs and people. A total of 6009 dogs were recorded, of which only 3.3% appeared to be unowned; unowned dogs had poorer welfare and were more likely to be wary of attempted approaches by people. The source of dogs, method of confinement used by owners, and whether dogs were sterilised differed between three breed types; purebred dogs, mixed breed, and Bali dogs (native breed). Three variables were found to have significant effects on the chances of not being vaccinated: age, dog type, and confinement. A mean of 3.81 roaming dogs per km of street surveyed was recorded along 28 sub-village routes. Responses to attitude statements showed that most people had a positive attitude towards dogs and vaccination and did not agree with culling. Knowledge of appropriate bite treatment and symptoms of rabies in dogs was good.

Reducing zoonotic and internal parasite burdens in pigs using a pig confinement system.

AIM: This study was designed to validate the effectiveness of the pig confinement system (PCS) in reducing the prevalence of zoonotic and internal parasite burdens in pigs. MATERIALS AND METHODS: Ten PCS households were selected together with 10 households practising traditional scavenging systems. Five pigs were monitored per household every 3 months for 15 months and blood and feces collected. Pigs received a single dose of oxfendazole at 30 mg/kg at baseline. Qualitative fecal examinations for intestinal parasite stages were performed, and serum was tested for antibodies to cysticercus of Taenia solium, Trichinella spp., and Toxoplasma gondii. RESULTS: Based on fecal examination, the prevalence of pigs positive for parasite eggs was reduced in PCS pigs over consecutive samplings (Ascaris suum [14.3% to 0%], Trichuris suis [46.9% to 8.3%], Strongyle-type eggs [81.6% to 8.3%], Physocephalus spp. [6.1% to 0%], and Metastrongylus apri [20.8% to 0%]) compared with increases in the number of pigs positive for parasite eggs in non-PCS pigs (T. suis [20-61.5%], Strongyle-type [60.4-80.8%], Physocephalus spp. [8.3-15.4%], and M. apri [20.8-34.6%]) and little change in pigs positive for A. suum (18.8-19.2%). While the prevalence of pigs with antibodies against to cysticerci of T. solium reduced in PCS pigs from 18% to 14%, the prevalence in non-PCS pigs increased from 42% to 52%. Antibodies to Trichinella were not detected, but the prevalence of T. gondii antibodies increased from 6% to 10% in PCS pigs and from 7% to 24% in non-PCS pigs. CONCLUSION: These data demonstrate the potential of a PCS to reduce the prevalence of pigs infected with zoonotic and internal parasites and thus the risk to human and pig health.